A number of different types of mass spectrometers are known to those skilled in the art, each having its own set of advantages and disadvantages. The present invention relates to methods of using the three-dimensional quadrupole ion trap mass spectrometer ("ion trap") which was initially patented in 1960 by Paul, et al. In recent years the ion trap mass spectrometer has grown in popularity in part due to its relatively low cost, ease of manufacture, and its unique ability to store ions over a large range of masses for relatively long periods of time. Nonetheless, the most common methods presently employed for using the ion trap do not yield very high mass resolution.
The quadrupole ion trap comprises a ring-shaped electrode and two end cap electrodes. Ideally, both the ring electrode and the end cap electrodes have hyperbolic surfaces that are coaxially aligned and symmetrically spaced. By placing a combination of AC and DC voltages (conventionally designated "V" and "U", respectively) on these electrodes, a quadrupole trapping field is created. This may be simply done by applying a fixed frequency (conventionally designated "f") AC voltage between the ring electrode and the end caps to create a quadrupole trapping field. The use of an additional DC voltage is optional, and in commercial embodiments of the ion trap no DC voltage is normally used. It can be shown that by using an AC voltage of proper frequency and amplitude, a wide range of masses can be simultaneously trapped.
The mathematics of the quadrupole trapping field created by the ion trap are well known and were described in the original Paul, et al., patent. For a trap having a ring electrode of a given radius equatorial r.sub.0, end caps displaced from the origin at the center of the trap along the axial line r=0 by a distance z.sub.0, and for given values of U, V and f, whether an ion of mass-to-charge ratio (m/e, also frequently designated m/z) will be trapped depends on the solution to the following two equations: ##EQU1##
Where .OMEGA. is equal to 2.pi.f.
Solving these equations yields values of a.sub.z and q.sub.z for a given ion species having the selected m/e. If the point (a.sub.z,q.sub.z) maps inside the stability envelop, the ion will be trapped by the quadrupole field. If the point (a.sub.z,q.sub.z) falls outside the stability envelop, the ion 7ill not be trapped and any such ions that are created within the trap will be quickly ejected. By changing the values of U, V or f one can affect the stability of a particular ion species. Note that from Eq. 1, when U=0, (i.e., when no DC voltage is applied to the trap), a.sub.z =0.
The typical method of using an ion trap consists of applying voltages to the trap electrodes to establish a trapping field which will retain ions over a wide mass range, introducing a sample into the ion trap, ionizing the sample, and then scanning the contents of the trap so that the ions stored in the trap are ejected and detected in order of increasing mass. Typically, ions are ejected through perforations in one of the end cap electrodes and are detected with an electron multiplier.
A number of methods exist for ionizing sample molecules. Most commonly, sample molecules are introduced into the trap and an electron beam is turned on ionizing the sample within the trap volume. This is referred to as electron impact ionization or "EI". Alternatively, ions of a reagent compound can be created within or introduced into the ion trap to cause ionization of the sample. This technique is referred to as chemical ionization or "CI". Other methods of ionizing the sample, such as photoionization using a laser beam, are also known. For purposes of the present invention the specific ionization technique used to create ions is not important.
Once the ions are formed and stored in the trap a number of techniques are available for isolating specific ions of interest, and for conducting so-called (MS).sup.n experiments. In (MS).sup.n experiments an isolated ion or group of ions, called "parent" ions, are fragmented creating "daughter" ions, which may be detected themselves or fragmented to create "granddaughter" ions, etc. Isolation techniques involve manipulating the trapping voltage(s) and/or using supplemental voltages as described below.
No matter what type of experiment is conducted within the ion trap, ultimately there will be a need to determine what ions are present in the trap. As noted above, this generally involves scanning the trap so that ions are ejected and detected. U.S. Pat. No. 4,540,884 describes a technique for scanning one or more of the basic trapping parameters, i.e., U, V or f, to sequentially cause trapped ions to become unstable and leave the trap. Unstable ions tend to leave in the axial direction and can be detected using a number of techniques, for example, as mentioned above, a electron multiplier connected to standard electronic circuitry.
In the preferred method taught by the '884 patent, the DC voltage, U, is set at 0. As noted, from Eq. 1 when U=0, then a.sub.z =0 for all mass values. As can be seen from Eq. 2, the value of q.sub.z is directly proportional to V and inversely proportional to the mass of the particle. Likewise, the higher the value of V the higher the value of q.sub.z. In the preferred embodiment the scanning technique of the '884 patent is implemented by ramping the value of V. As V is increased positively, the value of q.sub.z for a particular mass to charge ratio increases to the point where it passes from a region of stability to one of instability. Consequently, the trajectories of ions of increasing mass to charge ratio become unstable sequentially, and are detected when they exit the ion trap.
According to another known method of scanning the contents of an ion trap, a supplemental AC voltage is applied across the end caps of the trap to create an oscillating dipole field supplemental to the quadrupole field. In this method, the supplemental AC voltage has a different frequency than the primary AC voltage V. The supplemental AC voltage can cause trapped ions of specific mass to resonate at their so-called "secular" frequency in the axial direction. When the secular frequency of an ion equals the frequency of the supplemental voltage, energy is efficiently absorbed by the ion. When enough energy is coupled into the ions of a specific mass in this manner, those ions are ejected from the trap in the axial direction and subsequently detected. The technique of using a supplemental dipole field to excite specific ion masses is called axial modulation. Furthermore, axial modulation can be used to eject unwanted ions from the trap, and in connection with (MS).sup.n experiments to cause ions in the trap to collide with a buffer gas and fragment.
The secular frequency of an ion of a particular mass in an ion trap depends on the magnitude of the fundamental trapping voltage V. Thus, there are two ways of bringing ions of differing masses into resonance with the supplemental AC voltage. One can ramp the frequency of the supplemental voltage in a fixed trapping field, or one can vary the magnitude V of the trapping field while holding the frequency of the supplemental voltage constant. Typically, when using axial modulation to scan an ion trap, the frequency of the supplemental AC voltage is held constant and V is ramped so that ions of successively higher mass are ejected. The advantage of ramping the value of V is that it is relatively simple to perform and provides better linearity than can be attained by changing the frequency of the supplemental voltage. This method of scanning the trap is herein called resonance ejection scanning.
Resonance ejection scanning of trapped ions provides better sensitivity than can be attained using the mass instability technique taught by the '884 patent and produces narrower, better defined peaks. In other words, this technique produces better overall mass resolution. Resonance ejection also substantially increases the ability to analyze ions over a greater mass range.
In commercial embodiments of the ion trap using resonance ejection as a scanning technique, the frequency of the supplemental AC voltage is set at approximately one half of the frequency of the AC trapping voltage. It can be shown that the relationship of the frequency of the trapping voltage and the supplemental voltage determines the value of q.sub.z (as defined in Eq. 2 above) of ions that are at resonance. Indeed, sometimes the supplemental voltage is characterized in terms of the value of q.sub.z at which it operates.
A significant limiting factor in achieving very high mass resolution from the ion trap is in the rate at which the contents of the trap are scanned. Typically, commercial ion traps are designed to scan at a fixed rate of 5555 atomic mass units (amu's) per second; (stated equivalently, this is a scan rate of 190 .mu.s per amu).
Commercially, almost all ion traps are sold in connection with gas chromatographs (GC's) which serve, essentially, as input filters to the ion traps. However, the flow from a GC is continuous, and a modern high resolution GC produces narrow peaks, sometimes lasting only a matter of seconds. In order to detect narrow peaks, it is necessary to perform at least one complete scan of the ion trap per second. This, in turn, dictates the use of a fast scan rate in order to cover a wide range of masses.
In this context, mass resolutions of approximately 2,000 were typically all that could be achieved using resonance ejection as described above. Recently several experiments have been reported wherein significant improvements to this mass resolution have been achieved. However, the techniques used in these experiments all have significant shortcomings.
Initially, mass resolution was improved by simply slowing the scan rate by a factor of 100, such that the time required to scan one amu was increased to approximately 18 ms. This was shown to improve mass resolution to 33,000, at mass 502.
Another experiment to improve mass resolution involved scanning the frequency of the supplemental dipole voltage rather than the magnitude of the trapping voltage. However, this is difficult to do over a large range of masses, and requires more complex electronics. Nonetheless, one experiment using this technique has obtained mass resolution in excess of 45,000 at m/z 502.
Yet another improvement in mass resolution was shown when the scan rate was slowed even further, by a factor of 333 (which was realized by a 2000 fold attenuation in the rate in connection with a six-fold extension of the mass range). In this experiment, a mass resolution of 1,130,000 was achieved for CsI at m/z 3510. The FWHM (full width at half maximum) of the resulting peak was 3.5 mamu. In referring to high mass resolution spectroscopy it is generally more informative to quote the peak width at half height (i.e., FWHM) rather than the resolution itself.
While some of these improvements have been quite dramatic, there are several problems associated with them. In many circumstances a very slow scan rate is impractical due to the need to scan a wide range of masses in a relatively short time slow scanning is relatively high in noise, however, when using slow scanning it is difficult to utilize the traditional approach to improving signal-to-noise ratio by averaging the results of several scans. In particular, instabilities in the mass axis over time causes the location of the mass peaks to drift over time, and the longer the time between scans the greater the problem. It is generally believed that this problem is attributable to instabilities in the electronics of the ion trap, principally the RF electronics. Moreover, space charge differences in the ion trap from one experiment to the next, or over the course of a single experiment, can contribute to mass axis instability.
As mass resolution increases, the accuracy of the mass determination becomes a more difficult problem. While it may be possible, using the above described techniques, to obtain a very narrow mass peak, determining the exact mass number of the highly resolved peak is a wholly different and quite difficult problem. One approach that has been used to solve this has been to introduce an internal standard, for calibration purposes, of a reference compound of known mass, for example, CsI. One reported experiment describes a method whereby CsI atoms are introduced from outside the trap using a solid probe. This method, however, has not been shown to be effective when using very high resolution techniques, e.g., when mass resolutions greater than 50,000 are involved. Moreover, in many instances, the reference ion will not be close in mass to sample ions of interest. In such instances, the relatively long time needed to scan the ion trap between the calibration mass and the sample ions of interest, in connection with the aforementioned mass axis instabilities, precludes this from being a useful technique for very high mass resolution. For example, if the sample ion has a mass-to-charge ratio of 414 and the reference ion has a mass-to-charge ratio of 502 it would take approximately 18 seconds to scan between them at a scan rate of 5 amu/sec. While, in theory, it may be possible to try to select a calibration mass which is close in mass to the ion(s) of interest, so that the scanning time between them is minimized, this does not appear to be a practical solution.
In addition, as with most any instrument of its type, it is known that the dynamic range of an ion trap is limited, and that the most accurate and useful results are attained when the trap is filled with the optimal number of ions. If the too few ions are present in the trap, sensitivity is low and peaks may be overwhelmed by noise. If too many ions are present in the trap, space charge effects can significantly distort the trapping field, and peak resolution can suffer. The prior art has addressed this problem by using a so-called automatic gain control (AGC) technique whereby the total charge in the trap is integrated and held at a constant level. The AGC technique of the prior art does not distinguish how the total charge in the trap is distributed among the various masses, so that it does not determine whether the total integrated charge is distributed equally among all masses or if it resides at a single mass. In particular, the prior art AGC technique uses a fast "prescan" of the contents of the trap to integrate the charge present in the trap over the total mass range. While this approach is acceptable for normal low mass resolution scanning, at high resolution, it is extremely important to control the amount of charge due to ions having mass-to-charge ratios in the vicinity of a particular mass which is scanned at very high resolution.